Abstract
Vitamin D is an essential nutrient closely associated with the prevention of multiple diseases, including osteoporosis, diabetes, and cardiovascular disorders. The serum level of 25-hydroxyvitamin D [25(OH)D] is the primary biomarker for assessing vitamin D status, and its precise quantification is critical for clinical diagnosis and treatment. Chemiluminescence immunoassay (CLIA) and liquid chromatography–tandem mass spectrometry (LC-MS/MS) are widely used analytical methods; however, methodological discrepancies often lead to inconsistent results that may be further influenced by demographic factors such as age and gender. In this study, we evaluated the consistency and correlation between CLIA and LC-MS/MS measurements using a Generative Fuzzy Inference System (GENFIS). Analysis of 138 serum samples showed that LC-MS/MS produced significantly higher 25(OH)D concentrations than CLIA (p < 0.01; mean difference = 1.33 ± 3.71; 95% CI: − 5.95 to 8.61), though the two methods exhibited strong linear correlation and agreement (Cohen’s Kappa = 0.8257; R² = 0.9075; intraclass correlation coefficient = 0.93). GENFIS analysis indicated a possible 30–40-year female pattern of larger between the detection differences of two methods. To verify this finding, an additional 59 samples were analyzed, revealing a relative risk (RR) of 3.18 (95% CI: 1.71–5.93; p < 0.05) for this subgroup compared with other populations, supporting the GENFIS inference. These results highlight age- and gender-related differences in 25(OH)D measurement between CLIA and LC-MS/MS, providing valuable insights for improving the standardization and clinical interpretation of vitamin D testing.
Data availability
All data generated or analysed during this study are included in this published article and its supplementary information files.
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All authors have approved the manuscript and declare that it is not under consideration elsewhere. There are no conflicts of interest to disclose. The specific contributions of each author are as follows: Hongkun Liu served as the principal investigator and was responsible for study conception and manuscript drafting.Jianli Cui and Lei Zhang, as department professor, provided overall supervision, critical guidance, and final approval of the research.Kok Wai Wong and Sirui Li, professors and lecturers at the School of Information Technology, Murdoch University, advised on model architecture, refined the fuzzy-logic framework, and assisted in selecting appropriate AI methodologies.Hui Deng, Xinyi He, and Shuhao Liang managed sample acquisition, performed statistical analyses, and compiled the data.Yujie Li oversaw the quality control of the LC-MS/MS assays.Linlin Zhang contributed to manuscript revision and offered recommendations on experimental design.
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The study was reviewed and approved by the Ethics Committee of Sichuan Taikang Hospital (approval number: SCTK-IRB-2025-003). This research was conducted as a retrospective study using laboratory testing data from Sichuan Taikang Hospital. The Ethics Committee granted a waiver of informed consent in accordance with the Declaration of Helsinki and relevant national guidelines, as the study involved the analysis of anonymized data and posed no risk to participants. All methods were performed in accordance with the relevant guidelines and regulations.
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Liu, H., Li, S., Wong, K.W. et al. Analysis of chemiluminescence and liquid chromatography-mass spectrometry in 25-hydroxyvitamin D detection using fuzzy logic. Sci Rep (2026). https://doi.org/10.1038/s41598-026-41793-9
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DOI: https://doi.org/10.1038/s41598-026-41793-9
